This proposal will provide me with mentoring, hands on research experience and needed didactic training in order to develop into an independent physician-scientist. I am trained in Infectious Diseases and this proposal will advance my research interest in Hepatitis C virus (HCV). Steatosis, or fat accumulation within hepatocytes, is a major component of HCV-induced changes in liver histology. HCV genotype 3, unlike other genotypes, is an independent risk factor for both the presence and severity of steatosis. My preliminary sequence analysis of patient derived HCV RNA identified amino acid polymorphisms within Domain 3 of the Core protein that correlate with the presence or absence of steatosis. In vitro expression of these patient derived clones leads to intracellular fat accumulation. In this proposal, I will test the hypotheses that: 1) specific amino acid substitutions within Domain 3 of the Core protein are observed in patients with steatosis in HCV genotype 3 infection;2) these amino acid substitutions are required to cause intracellular lipid accumulation when introduced into cultured hepatocyte cell lines;3) intracellular fat in HCV infection contributes to fibrosis formation in the liver via interactions between HCV Core protein, Microsomal Triglyceride Transfer Protein (MTP), CD1d and NKT cells. In order to test the hypotheses listed above, I will use a combination of RT-PCR amplification of viral genes, in vitro expression, light and fluoresecent microscopy, recomibinant adenovirus infections in mice and flow cytometry. I will be using patient samples from a well characterized specimen bank from over 400 patients with HCV infection in a laboratory that focuses on basic liver biochemistry, molecular biology and intracellular signaling. I have assembled a team of mentors and consultants with expertise in liver biology and Hepatitis C clinical and molecular virology. We are examining differences between HCV genotype 3 isolates rather than between different genotypes. These stuides will provide a basis for studies of mechanisms whereby HCV infection causes intrahepatic fat accumulation that leads to fibrosis formation when no mechanism has previously been proposed. Delineation of the mechanisms underlying steatosis in HCV will lead to a greater insight into fibrosis and cancer formation. Results from this work may improve counselling of HCV patients with steatosis and may facilitate efforts to identify novel targets for anti-viral development.